Expression of genes related to muscle plasticity after strength and power training regimens

The purpose of our study was to compare the effects of 8-week progressive strength and power training regimens on strength gains and muscle plasticity [muscle fiber hypertrophy and phenotype shift, mammalian target of rapamycin (mTOR), regulatory-associated protein of mTOR (RAPTOR), rapamycin-insensitive companion of m-TOR (RICTOR), calcineurin and calcipressin gene expression]. Twenty-nine physically active subjects were divided into three groups: strength training (ST), power training (PT) and control (C). Squat 1 RM and muscle biopsies were obtained before and after the training period. Strength increased similarly for both ST and PT groups (P < 0.001). Fiber types I, IIa and IIb presented hypertrophy main time effect (P < 0.05). Only type IIb percentage decreased from pre- to post-test (main time effect, P < 0.05). mTOR and RICTOR mRNA expression increased similarly from pre- to post-test (P < 0.01). RAPTOR increased after training for both groups (P < 0.0001), but to a greater extent in the ST (P < 0.001) than in the PT group. 4EBP-1 decreased after training when the ST and PT groups were pooled (P < 0.05). Calcineurin levels did not change after training, while calcipressin increased similarly from pre- to post-test (P < 0.01). In conclusion, our data indicate that these training regimens produce similar performance improvements; however, there was a trend toward greater hypertrophy-related gene expression and muscle fiber hypertrophy in the ST group.

Resistance Training with Vascular Occlusion in Inclusion Body Myositis: A Case Study

GUALANO, B., M. NEVES JR, F. R. LIMA, A. L. PINTO, G. LAURENTINO, C. BORGES, L. BAPTISTA, G. G. ARTIOLI, M. S. AOKI, A. MORISCOT, A. H. LANCHA JR, E. BONFA, and C. UGRINOWITSCH. Resistance Training with Vascular Occlusion in Inclusion Body Myositis: A Case Study. Med Sci. Spot-is Exerc., Vol. 42, No. 2, pp. 250-254, 2010. Inclusion body myositis (IBM) is a rare idiopathic inflammatory myopathy that produces remarkable muscle weakness. Resistance training with vascular occlusion has been shown to improve muscle strength and cross-sectional area in other muscle wasting conditions. Purpose: We evaluated the efficacy of a moderate-intensity resistance training program combined with vascular occlusion by examining functional capacity, muscle morphology, and changes in the expression of genes related to muscle protein synthesis and proteolysis in a patient with IBM. Methods: A 65-yr-old man with IBM resistant to all proposed treatments underwent resistance training with vascular occlusion for 12 wk. Leg press one-repetition maximum; thigh cross-sectional area; balance, mobility, and muscle function; quality of life; and blood markers of inflammation and muscle damage were assessed at baseline and after the 12-wk program. The messenger RNA (mRNA) expression levels of mechanogrowth factor, mammalian target of rapamycin, atrogin-1, and muscle RING finger-1 were also quantified. Results: After the 12-wk training program, the patient`s leg press one-repetition maximum, balance and mobility function, and thigh cross-sectional area increased 15.9%, 60%, and 4.7%, respectively. All Short Form-36 Health Survey Questionnaire subscales demonstrated improvements as well, varying from 18% to 600%. mRNA expression of mechanogrowth factor increased 3.97-fold, whereas that of atrogin-1 decreased 0.62-fold. Muscle RING finger-1 and mammalian target of rapamycin mRNA levels were only slightly altered, 1.18- and 1.28-fold, respectively. Importantly, the exercise did not induce disease flare. Conclusions: We describe a novel, and likely the first, nonpharmacological therapeutic tool that might be able to counteract the muscle atrophy and the declining strength that usually occur in IBM.

Effect of eccentric exercise velocity on akt/mtor/p70(s6k) signaling in human skeletal muscle

It has been suggested that muscle tension plays a major role in the activation of intracellular pathways for skeletal muscle hypertrophy via an increase in mechano growth factor (MGF) and other downstream targets. Eccentric exercise (EE) imposes a greater amount of tension on the active muscle. In particular, high-speed EE seems to exert an additional effect on muscle tension and, thus, on muscle hypertrophy. However, little is known about the effect of EE velocity on hypertrophy signaling. This study investigated the effect of acute EE-velocity manipulation on the Akt/mTORCI/p70(S6K) hypertrophy pathway. Twenty subjects were assigned to either a slow (20 degrees.s(-1); ES) or fast EE (210 degrees.s(-1); EF) group. Biopsies were taken from vastus lateralis at baseline (B), immediately after (T1), and 2 h after (T2) the completion of 5 sets of 8 repetitions of eccentric knee extensions. Akt, mTOR, and p70(S6K) total protein were similar between groups, and did not change postintervention. Further, Akt and p70(S6K) protein phosphorylation were higher at T2 than at B for ES and EF. MGF messenger RNA was similar between groups, and only significantly higher at T2 than at B in ES. The acute manipulation of EE velocity does not seem to differently influence intracellular hypertrophy signaling through the Akt/mTORCI/p70S6K pathway.

Morphological and molecular characterization of cyanobacteria from a Brazilian facultative wastewater stabilization pond and evaluation of microcystin production

The cyanobacterial population in the Cajati waste stabilization pond system (WSP) from Sao Paulo State, Brazil was assessed by cell isolation and direct microscope counting techniques. Ten strains, belonging to five genera (Synechococcus, Merismopedia, Leptolyngbya, Limnothrix, and Nostoc), were isolated and identified by morphological and molecular analyses. Morphological identification of the isolated strains was congruent with their phylogenetic analyses based on 16S rDNA gene sequences. Six cyanobacterial genera (Synechocystis, Aphanocapsa, Merismopedia, Lyngbya, Phormidium, and Pseudanabaena) were identified by direct microscope inspection. Both techniques were complementary, since, of the six genera identified by direct microscopic inspection, only Merismopedia was isolated, and the four other isolated genera were not detected by direct inspection. Direct microscope counting of preserved cells showed that cyanobacteria were the dominant members (> 90%) of the phytoplankton community during both periods evaluated (summer and autumn). ELISA tests specific for hepatotoxicmicrocystins gave positive results for six strains (Synechococcus CENA108, Merismopedia CENA106, Leptolyngbya CENA103, Leptolyngbya CENA112, Limnothrix CENA109, and Limnothrix CENA110), and for wastewater samples collected from raw influent (3.70 mu g microcystins/l) and treated effluent (3.74 mu g microcystins/l) in summer. Our findings indicate that toxic cyanobacteria in WSP systems are of concern, since the treated effluent containing cyanotoxins will be discharged into rivers, irrigation channels, estuaries, or reservoirs, and can affect human and animal health.

Rotifer abundance, biomass, and secondary production after the recovery of hydrologic connectivity between a river and two marginal lakes (Sao Paulo, Brazil)

Rotifera density, biomass, and secondary production on two marginal lakes of Paranapanema River were compared after the recovery of hydrologic connectivity with the river (Sao Paulo State, Brazil). Daily samplings were performed in limnetic zone of both lakes during the rainy season immediately after lateral inflow of water and, in the dry period, six months after hydrologic connectivity recovery. In order to identify the factors that affect rotifer population dynamics, lake water level, volume, depth, temperature, transparency, dissolved oxygen, pH, alkalinity, conductivity, suspended solids, nutrients, and chlorophyll-a were determined. Variations of water physical and chemical factors that affect rotifer population were related to the lake-river degree of connection and to water level rising after drought. The water lateral inflow from the river resulted in an increase in lake water volume, depth, and transparency and a decrease in water pH, alkalinity, and suspended solids. The lake with the wider river connection, more frequent biota exchange, and larger amount of particulate and dissolved materials was richer and more diverse, while rotifer density, biomass, and productivity were lower in both periods studied. Density, biomass, and secondary production were higher in the lake with the smaller river connection and the higher physical and chemical stability. Our results show that the connectivity affects the limnological stability, associated to seasonality. Stable conditions, caused by low connectivity in dry periods, were related with high density, biomass and secondary production. Conversely, instability conditions in rainy periods were associated to elevated richness and diversity values, caused by exchange biota due to higher connectivity. (C) 2008 Elsevier GmbH. All rights reserved.

HEMOPLASMAS IN WILD CANIDS AND FEUDS IN BRAZIL

Hemotropic mycoplasmas, epicellular erythrocytic bacterial parasites lacking a cell wall, are the causative agents of infectious anemia in numerous mammalian species. The presence of hemotropic mycoplasmas in blood samples of neotropical and exotic wild canids and felids from Brazilian zoos were recorded using molecular techniques. Blood samples were collected from 146 Brazilian wild felids, 19 exotic felids, 3 European wolves (Canis lupus), and from 97 Brazilian wild canids from zoos in the Brazilian states of Sao Paulo and Mato Grosso and the Federal District. Using conventional polymerase chain reaction (PCR), this work found 22 (13%) wild felids positive to Candidatus Mycoplasma haemominutum [4 jaguars (Panthera onca); 3 pumas (Puma concolor); 10 ocelots (Leopardus pardalis); 2 jaguarondis (Puma yagouaroundi); and 3 little spotted cats (Leopardus tigrinus)]. Only one little spotted cat (Leopardus tigrinus) was positive to Mycoplasma haemofelis, and none was positive to Candidatus Mycoplasma turicensis. Two bush dogs (Speothos venaticus) were positive for a Mycoplasma sp. closely related to Candidatus Mycoplasma haematoparvum, and two European wolves were positive for a Mycoplasma sp. closely related to candidatus Mycoplasma haemominutum. This is the first study regarding the molecular detection of hemotropic mycoplasmas in wild canids.

Determination of Hg and diet identification in otter (Lontra longicaudis) feces

An analytical procedure for the determination of Hg in otter (Lontra longicaudis) feces was developed, to separate fish scales for the identification of the animal diet. Samples were washed with ultra-pure water and the suspension was sampled and transferred for digestion. The solubilization was performed with nitric-perchloric acid mixture, and detection carried out by the atomic fluorescence spectrometry (AFS). The quality of the analytical procedure was assessed by analyzing in-house standard solutions and certified reference materials. Total Hg concentrations were in the range of 7.6-156 ng g(-1) (July 2004), 25.6-277 ng g(-1) (January 2005) and 14.6-744 ng g(-1) (May 2005) that is approximately the same order of magnitude for all samples collected in two reservoirs at the Tiete River, Brazil. Although Hg concentrations varied with sampling periods and diet, high levels were correlated to the percentage of carnivorous fish scales present in the otter feces. (c) 2007 Elsevier Ltd. All rights reserved.

Effects of glutamine on the nuclear factor-kappaB signaling pathway of murine peritoneal macrophages

The aim of this study was to evaluate the effect of glutamine on the expression of proteins involved in the nuclear factor-kappaB (NF-kappa B) signaling pathway of murine peritoneal macrophages. Since glutamine is essential for the normal functioning of macrophages, it was hypothesized that in vitro glutamine supplementation would increase NF-kappa B activation. Peritoneal macrophages were pretreated with glutamine (0, 0.6, 2 and 10 mM) before incubation with lipopolysaccharide (LPS), and the effects of glutamine on the production of tumor necrosis factor-alpha and on the expression and activity of proteins involved in the NF-kappa B signaling pathway were studied by an enzyme linked immuno-sorbent assay, Western blotting, and an electrophoretic mobility shift assay. Glutamine treatment (2 and 10 mM) increased the activation of NF-kappa B in LPS-stimulated peritoneal macrophages (P < 0.05). In non-stimulated cells, glutamine treatment (2 and 10 mM) significantly reduced I kappa B-alpha protein expression (P < 0.05). Glutamine modulates NF-kappa B signaling pathway by reducing the level of I kappa B-alpha, leading to an increase in NF-kappa B within the nucleus in peritoneal macrophages.

Cyanobacteria and Cyanotoxin in the Billings Reservoir (Sao Paulo, SP, Brazil)

The Billings Complex and the Guarapiranga System are important strategic reservoirs for the city of Sao Paulo and surrounding areas because the water is used among other things, for the public water supply. They produce 19,000 liters of water per second and Supply water to 5.4 million people. Crude water is transferred from the Taquacetuba branch of the Billings Complex to the Guarapiranga Reservoir to regulate the water level of the reservoir. The objective of this study was to evaluate the water quality in the Taquacetuba branch, focusing on cyanobacteria and cyanotoxins. Surface water samples were collected in February (summer) and July (winter) of 2007. Analyses were conducted of physical, chemical, and biological variables of he water, cyanobacteria richness and density, and the presence of cyanotoxins. The water was classified as eutrophic-hypereutrophic. Cyanobacteria blooms were observed in both collection periods. The cyanobacteria bloom was most significant in July, reflecting lower water transparency and higher levels of total solids, suspended organic matter, chlorophyll-a, and cyanobacteria density in the surface water. Low richness and elevated dominance of the cyanobacteria were found in both periods. Cylindrospermopsis raciborskii was dominant in February, with 352 661.0 cel mL(-1), and Microcystis panniformis was dominant in July, with 1 866 725.0 cel mL(-1). Three variants of microcystin were found in February (MC-RR, MC-LR, MC-YR), as well as saxitoxin. The same variants of microcystin were found in July, but no saxitoxin was detected. Anatoxin-a and cylindropermopsin were not detected in either period. These findings are of great concern because the water in the Taquacetuba branch, which is transferred into the Guarapiranga Reservoir, is not treated nor managed. It is recommended that monitoring be intensified and more effective measures be taken by the responsible agencies to prevent the process of eutrophication and the consequent development of the cyanobacteria and their toxins.

Mitochondrial function in the yeast form of the pathogenic fungus Paracoccidioides brasiliensis

Differences between the respiratory chain of the fungus Paracoccidioides brasiliensis and its mammalian host are reported. Respiration, membrane potential, and oxidative phosphorylation in mitochondria from P. brasiliensis spheroplasts were evaluated in situ, and the presence of a complete (Complex I-V) functional respiratory chain was demonstrated. In succinate-energized mitochondria, ADP induced a transition from resting to phosphorylating respiration. The presence of an alternative NADH-ubiquinone oxidoreductase was indicated by: (i) the ability to oxidize exogenous NADH and (ii) the lack of sensitivity to rotenone and presence of sensitivity to flavone. Malate/NAD(+)-supported respiration suggested the presence of either a mitochondrial pyridine transporter or a glyoxylate pathway contributing to NADH and/or succinate production. Partial sensitivity of NADH/succinate-supported respiration to antimycin A and cyanide, as well as sensitivity to benzohydroxamic acids, suggested the presence of an alternative oxidase in the yeast form of the fungus. An increase in activity and gene expression of the alternative NADH dehydrogenase throughout the yeast`s exponential growth phase was observed. This increase was coupled with a decrease in Complex I activity and gene expression of its subunit 6. These results support the existence of alternative respiratory chain pathways in addition to Complex I, as well as the utilization of NADH-linked substrates by P. brasiliensis. These specific components of the respiratory chain could be useful for further research and development of pharmacological agents against the fungus.

Stereoselective analysis of thioridazine-2-sulfoxide and thioridazine-5-sulfoxide: An investigation of rac-thioridazine biotransformation by some endophytic fungi

The purpose of this study was to develop a method for the stereoselective analysis of thioridazine-2-sulfoxide (THD-2-SO) and thioridazine-5-sulfoxide (THD-5-SO) in culture medium and to study the biotransformation of rac-thioridazine (THD) by some endophytic fungi. The simultaneous resolution of THD-2-SO and THD-5-SO diastereoisomers was performed on a CHIRALPAK(R) AS column using a mobile phase of hexane: ethanol: methanol (92:6:2, v/v/v) + 0.5% diethylamine; UV detection was carried out at 262 nm. Diethyl ether was used as extractor solvent. The validated method was used to evaluate the biotransformation of THD by 12 endophytic fungi isolated from Tithonia diversifolia, Viguiera arenaria and Viguiera robusta. Among the 12 fungi evaluated, 4 of them deserve prominence for presenting an evidenced stereoselective biotransformation potential: Phomopsis sp. (TD2) presented greater mono-2-sulfoxidation to the form (S)-(SE) (12.1%); Glomerella cingulata (VA1) presented greater mono-5-sulfoxidation to the forms (S)-(SE) + (R)-(FE) (10.5%); Diaporthe phaseolorum (VR4) presented greater mono-2-sulfoxidation to the forms (S)-(SE) and (R)-(FE) (84.4% and 82.5%, respectively) and Aspergillus fumigatus (VR12) presented greater mono-2-sulfoxidation to the forms (S)-(SE) and (R)-(SE) (31.5% and 34.4%, respectively). (C) 2007 Elsevier B.V. All rights reserved.

Classical and alternative components of the mitochondrial respiratory chain in pathogenic fungi as potential therapeutic targets

The frequency of opportunistic fungal infection has increased drastically, mainly in patients who are immunocompromised due to organ transplant, leukemia or HIV infection. In spite of this, only a few classes of drugs with a limited array of targets, are available for antifungal therapy. Therefore, more specific and less toxic drugs with new molecular targets is desirable for the treatment of fungal infections. In this context, searching for differences between mitochondrial mammalian hosts and fungi in the classical and alternative components of the mitochondrial respiratory chain may provide new potential therapeutic targets for this purpose.

LC-MS-MS determination of ibuprofen, 2-hydroxyibuprofen enantiomers, and carboxyibuprofen stereoisomers for application in biotransformation studies employing endophytic fungi

The purpose of this study was the development and validation of an LC-MS-MS method for simultaneous analysis of ibuprofen (IBP), 2-hydroxyibuprofen (2-OH-IBP) enantiomers, and carboxyibuprofen (COOH-IBP) stereoisomers in fungi culture medium, to investigate the ability of some endophytic fungi to biotransform the chiral drug IBP into its metabolites. Resolution of IBP and the stereoisomers of its main metabolites was achieved by use of a Chiralpak AS-H column (150 x 4.6 mm, 5 mu m particle size), column temperature 8 degrees C, and the mobile phase hexane-isopropanol-trifluoroacetic acid (95: 5: 0.1, v/v) at a flow rate of 1.2 mL min(-1). Post-column infusion with 10 mmol L(-1) ammonium acetate in methanol at a flow rate of 0.3 mL min(-1) was performed to enhance MS detection (positive electrospray ionization). Liquid-liquid extraction was used for sample preparation with hexane-ethyl acetate (1:1, v/v) as extraction solvent. Linearity was obtained in the range 0.1-20 mu g mL(-1) for IBP, 0.05-7.5 mu g mL(-1) for each 2-OH-IBP enantiomer, and 0.025-5.0 mu g mL(-1) for each COOH-IBP stereoisomer (r >= 0.99). The coefficients of variation and relative errors obtained in precision and accuracy studies (within-day and between-day) were below 15%. The stability studies showed that the samples were stable (p > 0.05) during freeze and thaw cycles, short-term exposure to room temperature, storage at -20 degrees C, and biotransformation conditions. Among the six fungi studied, only the strains Nigrospora sphaerica (SS67) and Chaetomium globosum (VR10) biotransformed IBP enantioselectively, with greater formation of the metabolite (+)-(S)-2-OH-IBP. Formation of the COOH-IBP stereoisomers, which involves hydroxylation at C3 and further oxidation to form the carboxyl group, was not observed.

Enantioselective Analysis of Fluoxetine and Norfluoxetine by LC in Culture Medium for Application in Biotransformation Studies Employing Fungi

A liquid chromatography method is described for the analysis of fluoxetine and norfluoxetine enantiomers in fungi cultures. The analytes were separated simultaneously by LC employing a serial system. The resolution was performed using a mobile phase of ethanol: 15 mM ammonium acetate buffer solution, pH 5.9: acetonitrile (77.5:17.5:5, v/v/v). UV detection was at 227 nm. Hexane: isoamyl alcohol (98:2, v/v) was used as extractor solvent. The calibration curves were linear over the concentration range of 12.5-3,750 ng mL(-1) (r a parts per thousand yen 0.996). The values for intra- and inter-day precision and accuracy were a parts per thousand currency sign10% for all analytes. The validated method was used to evaluate fluoxetine biotransformation to its mammalian metabolite, norfluoxetine, by selected endophytic fungi. Although the desired biotransformation was not observed in the conditions used here, the method could be used to evaluate the biotransformation of fluoxetine by other fungi or to be extended to other matrices with adequate procedures for sample preparation.

Enantioselective analysis of propranolol and 4-hydroxypropranolol by CE with application to biotransformation studies employing endophytic fungi

A CE method is described for the enantioselective analysis of propranolol (Prop) and 4-hydroxypropranolol (4-OH-Prop) in liquid Czapek medium with application in the study of the enantioselective biotransformation of Prop by endophytic fungi. The electrophoretic conditions previously optimized were as follows: an uncoated fused-silica capillary, 4%w/v carboxymethyl-beta-CD in 25 mmol/L triethylamine/phosphoric acid (H(3)PO(4)) buffer at pH 9 as running electrolyte and 17 kV of voltage. UV detection was carried out at 208 nm. Liquid-liquid extraction using diethyl ether: ethyl acetate (1:1 v/v) as extractor solvent was employed for sample preparation. The calibration curves were linear over the concentration range of 0.25-10.0 mu g/mL for each 4-OH-Prop enantiomer and 0.10-10.0 mu g/mL for each Prop enantiomer (r >= 0.995). Within-day and between-day relative standard deviations and relative errors for precision and accuracy were lower than 15% for all the enantiomers. Finally, the validated method was used to evaluate Prop biotransformation in its mammalian metabolite 4-OH-Prop by some selected endophytic fungi. The screening of five strains of endophytic fungi was performed and all of them could biotransform Prop to some extent. Specifically, Glomerella cingulata (VA1) biotransformed 47.8% of (-)-(S)-Prop to (-)-(S)-4-OH-Prop with no formation of (+)-(R)4-OH-Prop in 72 h of incubation.